Daniel Day Lewis - My Beautiful Laundrette or A Room with a View, but I think many regard My Left Foot as his true breakthrough?

Colin Firth - Another Country, but I think Pride and Prejudice is the real answer
Helena Bonham Carter - A Room With a View (I simply refuse to acknowledge Lady Jane)
Robert Shaw - From Russia with Love
Hugh Grant - Four Weddings and a Funeral
Edward Fox - The Day of the Jackal
Richard Harris - This Sporting Life

I can't make up my mind on Julie Christie. Billy Liar, Darling or Dr. Zhivago?

Though I love Tootsie I would hardly call it a career defining movie for Hoffman. Surely The Graduate, Midnight Cowboy or even Straw Dogs, Papillon, All the Presidents Men, and even popular box office hits like Kramer vs Kramer and Marathon Man would be better qualified as career defining.

OK, perhaps some of the movies above are before your time and Tootsie may be the first film you became aware of him, but that's not the same as career defining, as in a role that an actor performed that made both the public, critics and peers sit up and took notice of the actor.

That said, In terms of Hoffman's career, without a doubt that would certainly be The Graduate.

Meryl Streep - Sophie's choice
Although she had success prior to that, it still remains an acclaimed performance and allowed Meryl to stand on her own as a lead actress, as The Deer Hunter and Kramer Vs Kramer only gave her supporting roles.

Anne Hathaway - The devil wears prada. She did a lot of family friends films before, but this role was very grown up for her. This film really led to the roles she did in films such as Batman and of course her now oscar winning performance of Les Miserables. Maybe not really a defining for her career but it was a stepping stone in the roles she is doing right now

Mmmmmm...not sure I would class that as career defining in the true sense, considering it's a very recent movie and he has been making movies (on and off) for over 30 years.

Maybe we need to actually define what we really mean here by 'career defining' - for me it's a film in which an actors performance first made everyone sit up and take notice, the film that effectively made their career take off. In most cases for an actor those movies are often very obvious to everyone, but not always. I think some people here are choosing either the first movie they specifically saw them in, or their favorite movie of that actor.

As far as Rourke is concerned, surely movies like Rumble Fish, Diner, 9 and a Half Weeks, Angel Heart would be more appropriate?

Glenn Close - Dangerous Liaisons (I know Fatal Attraction came first, but I think her performance in DL was far better)

Charlize Theron - Monster

I'd disagree with that.
Fatal Attraction was one of the defining movies of the 80s and after watching it recently it still has all of its power. Glenn Close fought tooth and nail for that part and it really was the performance that put her on the map.
DL as good as it is has become one of those slightly "forgotten" movies.
If you get down to the basics of "career-defining" i suppose it's the role that changed everything for that actor and put them where they are now.
If Glenn Close hadn't done FA her career would've been very different. DL wouldn't have done the same for her career.